Jem 1400 system

The 1−PLL complex was assessed by TEM using the negative staining method with ammonium molybdate (3.0 wt%). A solution of the complex was cast on a Cu100P grid coated with collodion films (thickness: 30–40 nm) supported by a carbon layer (thickness: 10–15 nm). The grid was pre-treated with low-energy plasma (DII-29020HD, JEOL Ltd., Tokyo, Japan) to hydrophilize the surface. The extra solution was removed using filter paper and subsequently air-dried. TEM observations were performed using the JEM-1400 system (JEOL Ltd., Tokyo, Japan) with an accelerating voltage of 80 kV.

NP characteristics such as size (diameter mean), polydispersity index (PDI), and zeta potential were acquired using dynamic light scattering (DLS; Zetasizer Nano, Malvern Instruments, Southborough, MA, USA). Working conditions were set as follows: sample dilution 1:20 (v/v) in HPLC-grade water, pH = 5, and temperature 25 °C.
Nanoparticle morphology was observed using a transmission electron microscope (TEM; JEM-1400 system, JEOL, Peabody, MA, USA). NPs were stained using 2% uranyl acetate; for analysis, the nanoparticle suspension was placed on 400-mesh carbon-coated grids.
A colorimetric assay was used to quantify the PVA remaining after nanoparticle dialysis [11 (link)]. In short, NPs powder was resuspended in 2 mL of 0.5 N NaOH and heated for 15 min at 60 °C. The samples were neutralized using 0.9 mL of 1 N HCl, and distilled water was added until a final volume of 5 mL. In the next step, 1.5 mL of distilled water, 0.5 0.05 M/0.15 M I₂/KI, and 3 mL of 0.65 M boric acid completed a volume of 10 mL. After 15 min of incubation at room temperature, the samples were measured using a spectrophotometer (Fisher Scientific, Pittsburg, PA, USA) at 690 nm. The standard curve for PVA was obtained using the same protocol for a concentration of 2.5–60 µg/mL.